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Development of a high-throughput assay to measure measles neutralizing antibodies.
PLoS One 2019; 14(8):e0220780Plos

Abstract

Measles virus is highly infectious and remains a leading cause of vaccine preventable deaths in children. Neutralizing antibody responses elicited by measles virus infection or immunization are a serological correlate of protection. We describe a high-throughput neutralization assay to improve surveillance for measles immunity. Measles virus-antibody mixtures were incubated on Vero cell monolayers and 24 hours later cell-lysates harvested and subjected to one-step SYBR green RT-qPCR to amplify a target sequence within the measles virus nucleoprotein gene. Neutralization endpoint titers were interpolated to determine the dilution that inhibited the relative amplicon copy number by at least 90% compared to the mean signal obtained in virus control wells in the absence of serum. Anti-measles virus and anti-measles hemagglutinin antisera specifically neutralized measles virus in the microneutralization RT-qPCR assay while pre-immune sera and sera raised against other viruses did not. The microneutralization RT-qPCR assay obeyed the Percentage Law for measles virus inputs ranging from 100-5000 TCID50/well. The linear range of the assay corresponds to measles antibody concentrations of 30 to 3000 mIU/mL. Bland-Altman analysis and two-way analysis of variance demonstrated that results obtained using the microneutralization RT-qPCR assay were comparable to those obtained using a plaque reduction neutralization test and correctly identified human serum samples that were seropositive (95% and 100%, sensitivity and specificity, respectively). Furthermore, these comparisons suggest that a concentration of 300 mIU/mL may be a conservative cut-point to use to identify individuals likely to be protected against severe measles disease when the endpoint is based on 90% inhibition of virus replication. Measles virus microneutralization RT-qPCR is a rapid, sensitive, specific, and robust assay for detecting measles neutralizing antibodies that may help to improve immunization strategies nationally and achieve measles elimination globally.

Authors+Show Affiliations

Division of Viral Products, Office of Vaccines Research and Review, Center for Biologics Evaluation Research, Food and Drug Administration, Silver Spring, Maryland, United States of America.Division of Viral Products, Office of Vaccines Research and Review, Center for Biologics Evaluation Research, Food and Drug Administration, Silver Spring, Maryland, United States of America.International Vaccine Access Center, Bloomberg School of Public Health, Johns Hopkins University, Baltimore, Maryland, United States of America.Division of Viral Products, Office of Vaccines Research and Review, Center for Biologics Evaluation Research, Food and Drug Administration, Silver Spring, Maryland, United States of America.

Pub Type(s)

Journal Article

Language

eng

PubMed ID

31415584

Citation

Alvarado-Facundo, Esmeralda, et al. "Development of a High-throughput Assay to Measure Measles Neutralizing Antibodies." PloS One, vol. 14, no. 8, 2019, pp. e0220780.
Alvarado-Facundo E, Audet S, Moss WJ, et al. Development of a high-throughput assay to measure measles neutralizing antibodies. PLoS ONE. 2019;14(8):e0220780.
Alvarado-Facundo, E., Audet, S., Moss, W. J., & Beeler, J. A. (2019). Development of a high-throughput assay to measure measles neutralizing antibodies. PloS One, 14(8), pp. e0220780. doi:10.1371/journal.pone.0220780.
Alvarado-Facundo E, et al. Development of a High-throughput Assay to Measure Measles Neutralizing Antibodies. PLoS ONE. 2019;14(8):e0220780. PubMed PMID: 31415584.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Development of a high-throughput assay to measure measles neutralizing antibodies. AU - Alvarado-Facundo,Esmeralda, AU - Audet,Susette, AU - Moss,William J, AU - Beeler,Judy A, Y1 - 2019/08/15/ PY - 2019/03/08/received PY - 2019/07/23/accepted PY - 2019/8/16/entrez PY - 2019/8/16/pubmed PY - 2019/8/16/medline SP - e0220780 EP - e0220780 JF - PloS one JO - PLoS ONE VL - 14 IS - 8 N2 - Measles virus is highly infectious and remains a leading cause of vaccine preventable deaths in children. Neutralizing antibody responses elicited by measles virus infection or immunization are a serological correlate of protection. We describe a high-throughput neutralization assay to improve surveillance for measles immunity. Measles virus-antibody mixtures were incubated on Vero cell monolayers and 24 hours later cell-lysates harvested and subjected to one-step SYBR green RT-qPCR to amplify a target sequence within the measles virus nucleoprotein gene. Neutralization endpoint titers were interpolated to determine the dilution that inhibited the relative amplicon copy number by at least 90% compared to the mean signal obtained in virus control wells in the absence of serum. Anti-measles virus and anti-measles hemagglutinin antisera specifically neutralized measles virus in the microneutralization RT-qPCR assay while pre-immune sera and sera raised against other viruses did not. The microneutralization RT-qPCR assay obeyed the Percentage Law for measles virus inputs ranging from 100-5000 TCID50/well. The linear range of the assay corresponds to measles antibody concentrations of 30 to 3000 mIU/mL. Bland-Altman analysis and two-way analysis of variance demonstrated that results obtained using the microneutralization RT-qPCR assay were comparable to those obtained using a plaque reduction neutralization test and correctly identified human serum samples that were seropositive (95% and 100%, sensitivity and specificity, respectively). Furthermore, these comparisons suggest that a concentration of 300 mIU/mL may be a conservative cut-point to use to identify individuals likely to be protected against severe measles disease when the endpoint is based on 90% inhibition of virus replication. Measles virus microneutralization RT-qPCR is a rapid, sensitive, specific, and robust assay for detecting measles neutralizing antibodies that may help to improve immunization strategies nationally and achieve measles elimination globally. SN - 1932-6203 UR - https://www.unboundmedicine.com/medline/citation/31415584/Development_of_a_high-throughput_assay_to_measure_measles_neutralizing_antibodies L2 - http://dx.plos.org/10.1371/journal.pone.0220780 DB - PRIME DP - Unbound Medicine ER -